RNAi Type of gene regulation.
Involves trigger dsRNA (may be from virus)
In C. elegans: Potent, Specific, Systemic, Inheritable.
Can also be seen in mammalian cells.
miRSC recruits deadenylase to remove poly-A tail which stabilises
mRNA, preventing degradation. Reduced half-life andreduction in
translation initiation. In mammals this is coupledwith decapping.
siRNA Unit responsible for interference in the RNAi pathway
Argaunaute Catalytic components of RISC complex
Piwi domain Found in argonaute, responsible for SLICER activity.
Slicer Activity which results in mRNA degradation
- Dicer cleaves trigger dsRNA
- siRNAs obtained.
- siRNA stabilized by Argonaute proteins in RISC.
- Like miRNA, siRNAs scan the entire transcriptome (mRNAs).
- Binds to complimentary sequence.
- Piwi domain cleaves mRNA by Slicer activity.
- mRNA degraded.
MicroRNA May have multiple targets, usually control genenetworks.
For humans: often found in fragile sites and genomic regions
involved ion cancers.
Heterochronic Timing is off. Larval stage forever, or skipped, etc.
Lin-4 Encodes small RNAs with antisense complementarity to lin-14.
Lin-4 base pairs to 7 parts of lin-14.
Mismatches helps dodge slicing machinery.
Lin-4 mutants have weird timing.
P bodies Store mRNA, dodge translation machinery.
Gel like. Biophysical properties favour interactions between RNA and
Have these activities:
- Decapping & mRNA degradation
- Storing mRNA until translation
- Aid in translational repression by miRNA miRNA cascade (ENDOGENOUS)
- Lin-4L precursor folds like a
hairpin, making dsRNA.
- Matures into lin-4S. Similar to an
- Lin-4 has partial complimentarity
with lin14 3’UTR.
- Prevents Slicer activity BUT
- Drosha cleaves pri-miRNA
transcript into pre-miRNA hairpin
- Exp5 brings it to cytoplasm
- Dicer matures it into siRNA duplex
by cleaving off hairpin
- siRNA bind Argo2, assembles into
- RISK scans mRNAs
miRNA mediated silencing.
No slicer activity!
Option 1: miRNAs prevent translational initiation
Option 2: miRNAs prevent PIC assembly
Option 3: (strongest)
- miRNAs bind at 3’, recruit Deadenylation complex.
- Poly-A tail removes
- mRNA is destabilized and degraded
- Reduction in translational initiation
- May be coupled to decapping
- mRNA is destabilized but NOT degraded!
o Controlled repression
The seed defines target for miRNA
Nucleotides 2 to 7 of miRNA
- Stronger base pairing brings RISK to UTR.
- 3’ UTRs do not have ORF, highly structured RNA balls.
- RISK may not get through UTR
Let-7 RAS regulation
Let-7 A microRNA. Can act as a tumor suppressor. Highly conserved from flies to humans.
Controls RAS expression in Human Cells.
Down-regulated in most tumors.
C. elegans let-7 mutants keep dividing, don’t differentiate.
miR-17-92 Oncogenic microRNA
Increased expression in lymphocytes linked with Lymphoproliferative
disease and autoimmunity in mice.
Let 7 seeds bind RAS UTRs
This relationship is highly conserved.
Let-7 lowers RAS expression in humans
Let-7 is downregulated in tumors, RAS goes up.
miRNAs and Cancer
miRNAs can be tumor suppressors or oncogenes.
**Recall: Tumor suppressor stops cancer genes, proto-oncogenes can become oncogenes. TS
repressive, Oncogene dominant.
c-Myc and miR-17~92
Polycistron. 4 families (mir17/mir20), (mir18), mir19, mir92. 6 miRNAs
Amplicon Causes genomic amplification, one source, many targets.
c-Myc Encodes a TF.
Cluster/Polycistron A group of microRNAs expressed from a single transcription unit.
Pri-mR-17~92 Has many pre-miRNAs attached, gives birth to 6 miRNAs.
Members have different targets that converge on thesame
miRNA family Share the same seed! Often leads to similar function.
miR-19 Oncogeniccomponent of mir-17~92
Selective pressure for it, amplified in many cancers.
Controls PTEN, more PTEN=Reduced apoptosis by reverting PI3K
miR-19 is a repressor of a repressor!
Other members control BIM o